Kovrigin Evgenii L, Loria J Patrick
Department of Chemistry, Yale University, P.O. Box 208107, New Haven, Connecticut 06520, USA.
J Am Chem Soc. 2006 Jun 21;128(24):7724-5. doi: 10.1021/ja061435a.
Through characterization of the solvent isotope effect on protein dynamics, we have examined determinants of the rate limitation to enzyme catalysis. A global conformational change in Ribonuclease A limits the overall rate of catalytic turnover. Here we show that this motion is sensitive to solvent deuterium content; the isotope effect is 2.2, a value equivalent to the isotope effect on the catalytic rate constant. We further demonstrate that the protein motion possesses a linear proton inventory plot, indicating that a single proton is transferred in the transition state. These results provide compelling evidence for close coupling between enzyme dynamics and function and demonstrate that characterization of the transition state for protein motion in atomic detail is experimentally accessible.
通过表征溶剂同位素对蛋白质动力学的影响，我们研究了酶催化速率限制的决定因素。核糖核酸酶A的全局构象变化限制了催化周转的总体速率。在此我们表明，这种运动对溶剂氘含量敏感；同位素效应为2.2，该值与对催化速率常数的同位素效应相当。我们进一步证明，蛋白质运动具有线性质子库存图，表明在过渡态中有单个质子转移。这些结果为酶动力学与功能之间的紧密耦合提供了令人信服的证据，并证明在原子细节上表征蛋白质运动的过渡态在实验上是可行的。
